Cyclic AMP-dependent phosphorylation of fructose-1,6-bisphosphatase in yeast

Purwin, Claudio; Leidig, Fritz; Holzer, Helmut

doi:10.1016/s0006-291x(82)80166-6

Cited by 104 publications

(48 citation statements)

References 14 publications

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“…Evidence has been provided that the rapid, reversible loss of about 60% of the fructose-1,6-diphosphatase 1 activity is mediated by phosphorylation of the enzyme catalyzed by a cAMP-dependent protein kinase (31,34). Considering this finding, it may be that the frucose-1,6-diphosphatase 1 activity changes during the cell cycle are regulated (at least in part) by a similar phosphorylationdephosphorylation mechanism as encountered previously for trehalase.…”

Section: Discussionmentioning

confidence: 72%

Changes in activities of several enzymes involved in carbohydrate metabolism during the cell cycle of Saccharomyces cerevisiae

et al. 1988

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Activity changes of a number of enzymes involved in carbohydrate metabolism were determined in cell extracts of fractionated exponential-phase populations of Saccharomyces cerevisiae grown under excess glucose. Cell-size fractionation was achieved by an improved centrifugal elutriation procedure. Evidence that the yeast populations had been fractionated according to age in the cell cycle was obtained by examining the various cell fractions for their volume distribution and their microscopic appearance and by flow cytometric analysis of the distribution patterns of cellular DNA and protein contents. Trehalase, hexokinase, pyruvate kinase, phosphofructokinase 1, and fructose-1,6-diphosphatase showed changes in specific activities throughout the cell cycle, whereas the specific activities of alcohol dehydrogenase and glucose-6-phosphate dehydrogenase remained constant. The basal trehalase activity increased substantially (about 20-fold) with bud emergence and decreased again in binucleated cells. However, when the enzyme was activated by pretreatment of the cell extracts with cyclic AMP-dependent protein kinase, no significant fluctuations in activity were seen. These observations strongly favor posttranslational modification through phosphorylation-dephosphorylation as the mechanism underlying the periodic changes in trehalase activity during the cell cycle. As observed for trehalase, the specific activities of hexokinase and phosphofructokinase 1 rose from the beginning of bud formation onward, finally leading to more than eightfold higher values at the end of the S phase. Subsequently, the enzyme activities dropped markedly at later stages of the cycle. Pyruvate kinase activity was relatively low during the Gl phase and the S phase, but increased dramatically (more than 50-fold) during G2. In contrast to the three glycolytic enzymes investigated, the highest specific activity of the gluconeogenic enzyme fructose-1,6-diphosphatase 1 was found in fractions enriched in either unbudded cells with a single nucleus or binucleated cells. The observed changes in enzyme activities most likely underlie pronounced alterations in carbohydrate metabolism during the cell cycle.Biochemical conversions which occur in bioreactors are the combined contributions of cells of different ages, i.e., cells at different stages of the cell cycle. Knowledge of possible variations in enzyme activities during the cell cycle is necessary to understand fully the metabolic behavior of the entire microbial culture. This requires methods to obtain subpopulations at defined stages of the cell cycle.In the past, the activity patterns of a variety of enzymes have been determined in synchronously dividing yeast cultures (10,27,28). However, as discussed by other investigators, many of these earlier results may have been flawed by technical artifacts, since the synchronization techniques that were used could cause serious perturbations in enzyme activity (6, 7,10,29). Creanor and Mitchison (7) developed a method of producing synchronous cultures of S...

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Section: Discussionmentioning

confidence: 72%

Changes in activities of several enzymes involved in carbohydrate metabolism during the cell cycle of Saccharomyces cerevisiae

et al. 1988

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“…Hence, covalent modification of phosphorylase involving the cAMP-mediated activation of an accessory kinase most likely occurs during adaptation to rich medium. Such conditions have been shown to promote increases in cAMP levels and cAMP-dependent kinase activity in yeasts (17,32,40,51). After glycogen depletion, subsequent exponential growth obviates any metabolic requirement to maintain phosphorylase in the cell.…”

Section: Resultsmentioning

confidence: 99%

Molecular analysis of GPH1, the gene encoding glycogen phosphorylase in Saccharomyces cerevisiae.

Hwang¹,

Tugendreich²,

Fletterick³

1989

Mol. Cell. Biol.

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In yeast cells, the activity of glycogen phosphorylase is regulated by cyclic AMP-mediated phosphorylation of the enzyme. We have previously cloned the gene for glycogen phosphorylase (GPH1) in Saccharomyces cerevisiae. To assess the role of glycogen and phosphorylase-catalyzed glycogenolysis in the yeast life cycle, yeast strains lacking a functional GPHI gene or containing multiple copies of the gene were constructed. GPHI was found not to be an essential gene in yeast cells. Haploid cells disrupted in GPHI lacked phosphorylase activity and attained higher levels of intracellular glycogen but otherwise were similar to wild-type cells. Diploid cells homozygous for the disruption were able to sporulate and give rise to viable ascospores. Absence of functional GPHI did not impair cells from synthesizing and storing trehalose. Increases in phosphorylase activity of 10-to 40-fold were detected in cells carrying multiple copies of GPHI-containing 2,um plasmid. Northern (RNA) analysis indicated that GPHI transcription was induced at the late exponential growth phase, almost simultaneous with the onset of intracellular glycogen accumulation. Thus, the low level of glycogen in exponential cells was not primarily maintained through regulating the phosphorylation state of a constitutive amount of phosphorylase. GPHI did not appear to be under formal glucose repression, since transcriptional induction occurred well in advance of glucose depletion from the medium.

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“…The cAMP level in yeast cells is known to be subject to rigorous feedback control. The glucose-triggered rapid rise in the intracellular cAMP level, which is dependent on Cdc25 and RAS (33,35), is immediately followed by return to the steady-state level (33)(34)(35)39; Fig. 7), and the return appears to require the activity of cAMP-dependent protein kinases (34).…”

Section: Discussionmentioning

confidence: 98%

“…By in vitro mutagenesis, we assigned a CAP-binding site of adenylyl cyclase to a small segment near its C terminus and created mutants which lost the ability to bind CAP. CAP binding was assessed first by observing the ability of the overproduced C-terminal 150 residues of adenylyl cyclase to sequester CAP, thereby suppressing the heat shock sensitivity of yeast cells bearing the activated RAS2 gene (33,35,39). A product of the CDC25 gene appears to convey the glucose signal to RAS proteins by promoting the GDP-GTP exchange on RAS (7,8,23,35).…”

mentioning

confidence: 99%

“…The yeast RAS proteins are essential controlling elements of adenylyl cyclase (6, 49), which catalyzes the production of cyclic AMP (cAMP), a second messenger critical for yeast cell growth (31). The RAS-adenylyl cyclase pathway has been implicated in transduction of a signal triggered by glucose to an intracellular environment where a protein phosphorylation cascade is induced by cAMP (33,35,39). A product of the CDC25 gene appears to convey the glucose signal to RAS proteins by promoting the GDP-GTP exchange on RAS (7,8,23,35).…”

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confidence: 99%

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Analysis of the function of the 70-kilodalton cyclase-associated protein (CAP) by using mutants of yeast adenylyl cyclase defective in CAP binding.

et al. 1993

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In Saccharomyces cerevisiae, adenylyl cyclase forms a complex with the 70-kDa cyclase-associated protein (CAP). By in vitro mutagenesis, we assigned a CAP-binding site of adenylyl cyclase to a small segment near its C terminus and created mutants which lost the ability to bind CAP. CAP binding was assessed first by observing the ability of the overproduced C-terminal 150 residues of adenylyl cyclase to sequester CAP, thereby suppressing the heat shock sensitivity of yeast cells bearing the activated RAS2 gene (33,35,39). A product of the CDC25 gene appears to convey the glucose signal to RAS proteins by promoting the GDP-GTP exchange on RAS (7,8,23,35). Yeast cells that possess activated RAS2 gene RAS2v"-"9 have an elevated level of intracellular cAMP and display abnormal phenotypes, including sensitivity to heat shock, sensitivity to nutritional starvation, and failure to sporulate (26,49).Yeast adenylyl cyclase, a product of the CYR1 gene (31), consists of 2,026 amino acid residues that comprise at least four domains: the N-terminal, the middle repetitive, the catalytic, and the C-terminal domains (24,53). The middle repetitive domain is composed of a repetition of 23-aminoacid leucine-rich motifs which have homology to the leucinerich repeat family proteins of yeasts, mammals, and Drosophila melanogaster (see references 15 and 52 and references therein). Studies using site-specific mutagenesis showed that any residue at particular positions of the consensus amino acids in the leucine-rich repeats was virtually unmutable without destruction of the interaction with RAS proteins (9,44,52). In addition, we recently identified a * Corresponding author.

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Cyclic AMP-dependent phosphorylation of fructose-1,6-bisphosphatase in yeast

Cited by 104 publications

References 14 publications

Changes in activities of several enzymes involved in carbohydrate metabolism during the cell cycle of Saccharomyces cerevisiae

Changes in activities of several enzymes involved in carbohydrate metabolism during the cell cycle of Saccharomyces cerevisiae

Molecular analysis of GPH1, the gene encoding glycogen phosphorylase in Saccharomyces cerevisiae.

Analysis of the function of the 70-kilodalton cyclase-associated protein (CAP) by using mutants of yeast adenylyl cyclase defective in CAP binding.

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